The present invention relates to self-assembled monolayers (SAMs) of alkanethiolates.
SAMs of alkanethiolates on gold are an important class of model substrates for mechanistic studies of the interactions of proteins and cells with surfaces. They have been used in monolayers that are inert in biological fluidsxe2x80x94in that they prevent protein adsorption and cell adhesionxe2x80x94and provide surfaces for patterning the positions and shapes of attached cells (Mrksich, M.; Dike, L. E.; Tien, J. Y.; Ingber, D. E.; Whitesides, G. M. Exp. Cell Res. 1997, 235, 305-313; and Chen, C. S.; Mrksich, M.; Huang, S.; Whitesides, G. M.; Ingber, D. E. Science 1997, 276, 1345-1347). The attachment of ligands to these inert SAMs gives surfaces to which proteins and other receptors selectively bind. Monolayers presenting peptide ligands, for example, have been used to control the adhesion of cells (Roberts, C.; Chen, C. S.; Mrksich, M.; Martichonok, V.; Ingber, D. E.; Whitesides, G. M. J. Am. Chem. Soc. 1998, 120, 6548-6555; and Houseman, B. T.; Mrksich, M. J. Org. Chem. 1998, 120, 6548-6555) and monolayers presenting oligonucleotides have been used for probing gene expression in cells (Bamdad, C. Biophys. J. 1998, 75, 1997-2003).
A new challenge in biointerfacial science is to design dynamic substrates that can alter, in real-time, the display of ligands, and hence interactions of proteins and cells with the substrate (Lau, A. N. K.; Miller, L. L. J. Am. Chem. Soc. 1983, 105, 5271-5277; and Ding, Z.; Long, C. J.; Hayashi, Y.; Bulmus, E. V.; Joffman, A. S.; Stayton, P. S. Bioconjugate Chem. 1999, 10, 395-400). Previously, a dynamic SAM that could be switched from a state that is initially inert to a state that permits the Diels-Alder mediated immobilization of ligands has been demonstrated, which in turn provides a strategy to activate the selective binding of proteins to a substrate (Yousaf, M. N.; Mrksich, M. J. Am. Chem. Soc. 1999, 121, 4286-4287). Previous examples of substrates that can release attached cells have relied on a thermally responsive poly(N-isopropylacrylamide) film (T. Okano, N. Yamata, J. Sakai, Y. Sakurai, Biomaterials 1995, 16, 297-303; and Y. Ito, G. Chen, Y. Guan, Y. Imanishi, Lanmuir 1997, 13, 2756-2759).
It would be desirable to have a dynamic monolayer that can selectively release immobilized ligands. Such a monolayer could be used to regulate, in real-time, the ligand-receptor interactions between a cell and the substrate to which it is attached, and give dynamic control of the proteins presented on the surface of a monolayer. It may also be used in microsystems to regulate the release of reagents or biomolecules from a solid or gel phase.
In a first aspect, the present invention is an alkanethiol of formula (1):
HSxe2x80x94Lxe2x80x94Q1xe2x80x94Txe2x80x94Q2xe2x80x94Mxe2x80x94Gxe2x80x94Zxe2x80x83xe2x80x83(1),
wherein xe2x80x94Lxe2x80x94 is xe2x80x94(ALjxe2x80x94BLkxe2x80x94ELlxe2x80x94DL)mxe2x80x94;
each AL, BL, EL and DL are individually C(RLARLAxe2x80x2)xe2x80x94, xe2x80x94C(RLB, RLBxe2x80x2)xe2x80x94, xe2x80x94C(RLERLExe2x80x2)xe2x80x94, and xe2x80x94C(RLDRLDxe2x80x2)xe2x80x94, respectively;
each RLA, RLB, RLE and RLD are individually H, or any two of RLA, RLB, RLE and RLD together form a bond, or RLA, RLB, RLE and RLD together with the atoms to which they are bonded form a six-membered aromatic ring;
each RLAxe2x80x2, RLBxe2x80x2, RLExe2x80x2 and RLDxe2x80x2 are individually H, or any two of RLAxe2x80x2, RLBxe2x80x2, RLExe2x80x2 and RLDxe2x80x2 together form a bond, or RLAxe2x80x2, RLBxe2x80x2, RLExe2x80x2 and RLDxe2x80x2 together with the atoms to which they are bonded form a six-membered aromatic ring;
each j, k and l are individually either 0 or 1;
m is 1 to 5;
xe2x80x94Q1xe2x80x94 and xe2x80x94Q2xe2x80x94 are each individually selected from the group consisting of 
xe2x80x94Txe2x80x94 is a moiety of formula (2) or formula (3) 
R1 and R2 are each individually selected from the group consisting of H and OH;
a is 0 to 3;
b is 0 to 3;
 indicates that the chirality of the carbon atom to which it is attached may be either R or S;
n is 1 to 6;
xe2x80x94Mxe2x80x94 is xe2x80x94(Axxe2x80x94Byxe2x80x94Ezxe2x80x94D)wxe2x80x94,
A is xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94N(RA)xe2x80x94 or xe2x80x94C(RARAxe2x80x2)xe2x80x94;
B is xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94N(RB)xe2x80x94 or xe2x80x94C(RBRBxe2x80x2)xe2x80x94;
E is xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94N(RE)xe2x80x94 or xe2x80x94C(RERExe2x80x2)xe2x80x94;
D is xe2x80x94C(RDRDxe2x80x2)xe2x80x94;
each RA, RB, RE and RD is selected from the group consisting of H, alkyl, alkenyl, alkynyl, aryl and heterocyclic radical, or any two of RA, RB, RE and RD together form a bond, or any two of RA, RB, RE and RD together with the atoms to which they are bonded for a ring;
each RAxe2x80x2, RBxe2x80x2, RExe2x80x2 and RDxe2x80x2 is selected from the group consisting of H, alkyl, alkenyl, alkynyl, aryl and heterocyclic radical, or any two of RAxe2x80x2, RBxe2x80x2, RExe2x80x2 and RDxe2x80x2 together form a bond, or any two RAxe2x80x2, RBxe2x80x2, RExe2x80x2 and RDxe2x80x2 together with the atoms to which they are bonded for a ring;
each x, y and z is either 0 or 1;
xe2x80x94Gxe2x80x94 is a moiety of formula (4) 
R3, R4, and R5 are each individually a xe2x80x94CH2xe2x80x94 group substituted with a hydrogen or an alkyl group;
R6 is a hydrogen, alkyl, alkenyl, alkynyl, aryl, halogen, heterocyclic radical or nitro group;
X is a bond or xe2x80x94(AXpxe2x80x94BXqxe2x80x94EXrxe2x80x94DX)sxe2x80x94;
each AX, BX, EX and DX is individually C(RXARXAxe2x80x2)xe2x80x94, xe2x80x94C(RXBRBxe2x80x2)xe2x80x94, xe2x80x94C(RXERExe2x80x2)xe2x80x94, or xe2x80x94C(RXDRXDxe2x80x2)xe2x80x94, respectively;
each RXA, RXB, RXE and RXD is individually H, or any two of RXA, RXB, RXE and RXD together form a bond, or RXA, RXB, RXE and RXD together with the atoms to which they are bonded form a six-membered aromatic ring;
each RXAxe2x80x2, RXBxe2x80x2, RXExe2x80x2 and RXDxe2x80x2 is individually H, or any two of RXAxe2x80x2, RXBxe2x80x2, RXExe2x80x2 and RXDxe2x80x2 together form a bond, or RXAxe2x80x2, RXBxe2x80x2, RXExe2x80x2 and RXDxe2x80x2 together with the atoms to which they are bonded form a six-membered aromatic ring;
each p, q and r is individually either 0 or 1;
s is 1 to 5;
Y is xe2x80x94Oxe2x80x94 or xe2x80x94NHxe2x80x94;
R7 and R8 are both single bonds, or together with the atoms to which they are bonded from one or more fused rings, substituted or unsubstituted, optionally containing heteroatoms; and
xe2x80x94Z is a leaving group.
In a second aspect, the present invention is a disulfide of formula (5):
Jxe2x80x94Sxe2x80x94Sxe2x80x94Lxe2x80x94Q1xe2x80x94Txe2x80x94Q2xe2x80x94Mxe2x80x94Gxe2x80x94Zxe2x80x83xe2x80x83(5),
where xe2x80x94Lxe2x80x94, xe2x80x94Q1xe2x80x94, xe2x80x94Q2xe2x80x94, xe2x80x94Txe2x80x94, xe2x80x94Mxe2x80x94, xe2x80x94Gxe2x80x94, and xe2x80x94Z have the same meaning as in formula (1),
xe2x80x94J is selected from the group consisting of H, halogen, R, xe2x80x94OR, xe2x80x94NRRxe2x80x2, xe2x80x94C(O)R, and xe2x80x94C(O)OR;
R is selected from the group consisting of alkyl, alkenyl, alkynyl, aryl and heterocyclic radical; and
Rxe2x80x2 is selected from the group consisting of H, alkyl, alkenyl, alkynyl, aryl and heterocyclic radical.
In a third aspect, the present invention is a substrate, comprising:
(i) a surface layer comprising gold, and
(ii) a plurality of moieties, on at least a portion of said surface layer,
wherein said moieties are alkanethiolate moieties of formula (8):
Surfxe2x80x94Sxe2x80x94Lxe2x80x94Q1xe2x80x94Txe2x80x94Q2xe2x80x94Mxe2x80x94Gxe2x80x94Zxe2x80x83xe2x80x83(8)
where xe2x80x94Lxe2x80x94, xe2x80x94Q1xe2x80x94, xe2x80x94Q2xe2x80x94, xe2x80x94Txe2x80x94, xe2x80x94Mxe2x80x94, xe2x80x94Gxe2x80x94, and xe2x80x94Z have the same meaning as in formula (1), and Surf designates where the moiety is attached to the surface.
In a fourth aspect, the present invention is a method of releasing a leaving group, comprising:
applying a reducing potential to a surface.
Definitions
xe2x80x9cAlkylxe2x80x9d (or alkyl- or alk-) refers to a substituted or unsubstituted, straight, branched or cyclic hydrocarbon chain, preferably containing from 1 to 20 carbon atoms. More preferred alkyl groups are alkyl groups containing from 7 to 16 carbon atoms. Preferred cycloalkyls have from 3 to 10, preferably 3-6, carbon atoms in their ring structure. Suitable examples of unsubstituted alkyl groups include methyl, ethyl, propyl, isopropyl, cyclopropyl, butyl, iso-butyl, tert-butyl, sec-butyl, cyclobutyl, pentyl, cyclopentyl, hexyl, cyclohexyl, and the like. xe2x80x9cAlkylarylxe2x80x9d and xe2x80x9calkylheterocyclicxe2x80x9d groups are alkyl groups covalently bonded to an aryl or heterocyclic group, respectively.
xe2x80x9cAlkenylxe2x80x9d refers to a substituted or unsubstituted, straight, branched or cyclic, unsaturated hydrocarbon chain that contains at least one double bond, and preferably 2 to 20, more preferably 7 to 16, carbon atoms. Exemplary unsubstituted alkenyl groups include ethenyl (or vinyl)(xe2x80x94CHxe2x95x90CH2), 1-propenyl, 2-propenyl (or allyl)(xe2x80x94CH2xe2x80x94CHxe2x95x90CH2), 1,3-butadienyl (xe2x80x94CHxe2x95x90CHCHxe2x95x90CH2), 1-butenyl (xe2x80x94CHxe2x95x90CHCH2CH3), hexenyl, pentenyl, 1,3,5-hexatrienyl, and the like. Preferred cycloalkenyl groups contain five to eight carbon atoms and at least one double bond. Examples of cycloalkenyl groups include cyclohexadienyl, cyclohexenyl, cyclopentenyl, cycloheptenyl, cyclooctenyl, cyclohexadienyl, cycloheptadienyl, cyclooctatrienyl and the like.
xe2x80x9cAlkynylxe2x80x9d refers to a substituted or unsubstituted, straight, branched or cyclic unsaturated hydrocarbon chain containing at least one triple bond, and preferably 2 to 20, more preferably 7 to 16, carbon atoms.
xe2x80x9cArylxe2x80x9d refers to any monovalent aromatic carbocyclic or heteroaromatic group, preferably of 3 to 10 carbon atoms. The aryl group can be monocyclic (i.e. phenyl (or Ph)) or polycyclic (i.e. naphthyl) and can be unsubstituted or substituted. Preferred aryl groups include phenyl, naphthyl, furyl, thienyl, pyridyl, indolyl, quinolinyl or isoquinolinyl.
xe2x80x9cHalogenxe2x80x9d (or halo-) refers to fluorine, chlorine, iodine or bromine. The preferred halogen is fluorine or chlorine.
xe2x80x9cHeterocyclic radicalxe2x80x9d refers to a stable, saturated, partially unsaturated, or aromatic ring, preferably containing 5 to 10, more preferably 5 or 6, atoms. The ring can be substituted 1 or more times (preferably 1, 2, 3, 4 or 5 times) with a substituent. The ring can be mono-, bi- or polycyclic. The heterocyclic group consists of carbon atoms and from 1 to 3 heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur. The heteroatoms can be protected or unprotected. Examples of useful heterocyclic groups include substituted or unsubstituted, protected or unprotected acridine, benzathiazoline, benzimidazole, benzofuran, benzothiophene, benzothiazole, benzothiophenyl, carbazole, cinnoline, furan, imidazole, 1H-indazole, indole, isoindole, isoquinoline, isothiazole, morpholine, oxazole (i.e. 1,2,3-oxadiazole), phenazine, phenothiazine, phenoxazine, phthalazine, piperazine, pyridine, purine, pyrazine, pyrazole, pyridazine, pyridine, pyrimidine, pyrrole, quinazoline, quinoline, quinoxaline, thiazole, 1,3,4-thiadiazole, thiophene, 1,3,5-triazines, triazole (i.e. 1,2,3-triazole), and the like.
xe2x80x9cSubstitutedxe2x80x9d means that the moiety contains at least one, preferably 1-3 substituent(s). Suitable substituents include hydrogen (H) and hydroxyl (xe2x80x94OH), amino (xe2x80x94NH2), oxy (xe2x80x94Oxe2x80x94), carbonyl (xe2x80x94COxe2x80x94), thiol, alkyl, alkenyl, alkynyl, alkoxy, halo, nitrile, nitro, aryl and heterocyclic groups. These substituents can optionally be further substituted with 1-3 substituents. Examples of substituted substituents include carboxamide, alkylmercapto, alkylsulphonyl, alkylamino, dialkylamino, carboxylate, alkoxycarbonyl, alkylaryl, aralkyl, alkylheterocyclic, and the like.
xe2x80x9cDisulfidexe2x80x9d means a compound containing a bond between two sulfur atoms.
xe2x80x9cAlkanethiolxe2x80x9d means a compound containing an alkyl group bonded to an SH group.
xe2x80x9cAlkanethiolatexe2x80x9d means a moiety corresponding to an alkanethiol without the hydrogen of the SH group.
xe2x80x9cAlkylenexe2x80x9d refers to a substituted or unsubstituted, straight, branched or cyclic hydrocarbon chain, preferably containing from 1 to 20 carbon atoms. More preferred alkylene groups are lower alkylene groups, i.e., alkylene groups containing from 1 to 6 carbon atoms. Preferred cycloalkylenes have from 3 to 10, preferably 3-6, carbon atoms in their ring structure. Suitable examples of unsubstituted alkylene groups include methylene, xe2x80x94(CH2)nxe2x80x94, xe2x80x94CH2xe2x80x94CH(CH3)xe2x80x94, xe2x80x94(C6H10)xe2x80x94 where the carbon atoms form a six-membered ring, and the like.
xe2x80x9cPolypeptidexe2x80x9d refers to a molecule or moiety containing two or more amino acids bound through a peptide linkage. Examples include proteins such as antibodies, enzymes, lectins and receptors. Preferred proteins include avidin and streptavidin.
xe2x80x9cPolynucleotidexe2x80x9d refers to a molecule or moiety containing two or more nucleic acid such as single or double stranded RND, DNA and PNA (protein nucleic acid).
xe2x80x9cCarbohydratexe2x80x9d refers to a molecule or moiety that contains one or more sugars, such as mannose, sucrose, glucose, cellulose, chitin, and chitosan.
xe2x80x9cLigandxe2x80x9d refers to a binding partner of a receptor. Examples include cytokines, and chemokines.
xe2x80x9cHaptenxe2x80x9d refers to a molecule or moiety that will bind to an antibody specific for that hapten. Examples include fluorescein, and phosphotyrosine.
All other acronyms and abbreviations have the corresponding meaning as published in journals relative to the art of chemistry.